1. Field of the Invention
The present invention relates to methods and devices for the analysis of cell samples where the samples are pure populations or subpopulations of desired types. In particular, the present invention allows for direct comparison of protein content and protein characteristics between proteins isolated from tumor and normal cells from the same tissue sample.
2. Discussion of the Background
Proteomics is the study of variations in cellular protein levels between normal and disease states. As the field of molecular biology moves beyond genomics to proteomics, there is a growing need for a direct method to monitor the levels of expressed proteins in developing, diseased or genetically altered tissues. Direct monitoring of tissues is difficult because of their heterologous, three-dimensional structure. This structure is the result of the strong adhesive interaction of the various cell types with adjacent cells, connective stroma, blood vessels, glandular and muscle components, adipose cells, and inflammatory or immune cells. The amount and type of protein expressed by cells in such a native tissue environment may be quite different from that of more easily studied cultured or transplanted cells. This consideration requires a direct means of measuring protein levels to obtain results reflecting in vivo conditions.
Previous methods for extracting and analyzing protein macromolecules from tissue subpopulations have included UV laser ablation of unwanted tissue regions (Meirer-Ruge et al., The laser in the Lowry technique for microdissection of freeze-dried tissue slices, 8 Histochemical J. 384 (1976)) and oil well isolation of tissue cells (Matschinsky et al., Quantitative histochemical analysis of glycolytic intermediates and cofactors with an oil well technique, 16 (1) J. Histochemical Cytochem. 29 (1968)). These methods were complicated, labor intensive, and did not utilize protein stabilizers.
Because of these limitations, a number of direct comparisons between tumor cells and normal cells have not been achievable. There has been no way to directly compare, without the danger of cross-contamination, the spectrum of proteins contained in normal cells with the proteins in tumor cells in a single tissue. Many hypotheses about altered protein levels in tumor cells have been based on work on cells lines, and the ability for continued growth in culture by the cell line injects yet another variable into the results. There has also been no way to directly quantify differences in protein amount between tumor and normal tissue. Much of the work in this area has been done using immunohistochemistry, imparting the limitations as to quantification and specificity discussed above. Moreover, there has been no way to compare the protein content of various stages of tumor development, or to compare the characteristics of proteins found in tumor cells to those in normal cells. Nor could a protein's amino acid sequence or binding characteristics be performed. And, because such comparative protein studies could not be done, it was difficult to reliably determine the source of a tumor metastasis, unless the probable tumor source was already known.